The specific aims of this proposal are to gain a molecular understanding of how the heat shock transcription factor is activated by heat shock, and to determine how the activated factor stimulates transcription of heat shock genes. In addition we would like to understand the mechanism by which heat shock shuts down the transcription of other genes. We plan on performing a systematic and comprehensive molecular analysis of the regulation and activities of the S. cerevisiae and Drosophila heat shock transcription factors. The regulatory domains of the S. cerevisiae HSF have been studied in detail. in my lab, and it is proposed to continue and further refine these studies. We have also examined the biochemical properties of the Drosophila HSTF and, having cloned the gene for the Drosophila HSF, we wish now to compare and contrast the modes of regulation of the Drosophila factor with that of S. cerevisiae. While we were able to use yeast genetics to examine the regulatory domains of the S. cerevisiae HSTF we do not have this capacity in Drosophila. To study the Drosophila factor we will use a combination of transient transfection of cultured cells as well as stable lines and transgenic flies. With these systems we should be able to dissect the different modes of regulation of the yeast and Drosophila HSTFs and determine the domains of the proteins that are involved in this regulation. Detailed studies on the mechanism of transcriptional activation are proposed which include in vitro analysis of the HSTFs in reconstituted systems. A long term goal is to determine the structure of the S. cerevisiae and Drosophila HSTFs in collaboration with Dr. Doug Rees at Caltech.